pRS405

Yeast integrative vector with a LEU2 marker and an MCS derived from pBLUESCRIPT II.
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AatII (5442) ZraI (5440) ScaI (5000) BsaI (4581) AhdI (4520) AlwNI (4043) PspFI (3935) BseYI (3931) BspQI - SapI (3511) lac operator SacI (3233) Eco53kI (3231) SacII (3224) AleI (3223) EagI - NotI (3212) XbaI (3205) SpeI (3199) BamHI (3193) SmaI (3189) TspMI - XmaI (3187) PstI (3185) HindIII (3163) SalI (3148) AbsI - PaeR7I - PspXI - XhoI (3142) ApaI (3137) PspOMI (3133) NaeI (2801) NgoMIV (2799) PfoI (46) PflFI - Tth111I (192) BsrGI (632) BfuAI - BspMI (759) XcmI (1201) BbsI (1301) AflII (1438) AgeI (1503) BstEII (1718) PpuMI (1875) HpaI (2174) KasI (2203) NarI (2204) SfoI (2205) PluTI (2207) BsaAI - DraIII (2698) BtgZI (2699) pRS405 5507 bp
AatII  (5442)
1 site
G A C G T C C T G C A G
ZraI  (5440)
1 site
G A C G T C C T G C A G
ScaI  (5000)
1 site
A G T A C T T C A T G A
BsaI  (4581)
1 site
G G T C T C N C C A G A G N ( N ) 4

Sticky ends from different BsaI sites may not be compatible.
BsaI can be used between 37°C and 50°C.
AhdI  (4520)
1 site
G A C N N N N N G T C C T G N N N N N C A G

The 1-base overhangs produced by AhdI may be hard to ligate.
Sticky ends from different AhdI sites may not be compatible.
AlwNI  (4043)
1 site
C A G N N N C T G G T C N N N G A C

Sticky ends from different AlwNI sites may not be compatible.
PspFI  (3935)
1 site
C C C A G C G G G T C G
BseYI  (3931)
1 site
C C C A G C G G G T C G

After cleavage, BseYI can remain bound to DNA and alter its electrophoretic mobility.
BspQI  (3511)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different BspQI sites may not be compatible.
SapI  (3511)
1 site
G C T C T T C N C G A G A A G N N N N

Sticky ends from different SapI sites may not be compatible.
SapI gradually settles in solution, so a tube of SapI should be mixed before removing an aliquot.
SacI  (3233)
1 site
G A G C T C C T C G A G
Eco53kI  (3231)
1 site
G A G C T C C T C G A G
SacII  (3224)
1 site
C C G C G G G G C G C C

Efficient cleavage requires at least two copies of the SacII recognition sequence.
AleI  (3223)
1 site
C A C N N N N G T G G T G N N N N C A C
EagI  (3212)
1 site
C G G C C G G C C G G C
NotI  (3212)
1 site
G C G G C C G C C G C C G G C G
XbaI  (3205)
1 site
T C T A G A A G A T C T
SpeI  (3199)
1 site
A C T A G T T G A T C A
BamHI  (3193)
1 site
G G A T C C C C T A G G

After cleavage, BamHI-HF® (but not the original BamHI) can remain bound to DNA and alter its electrophoretic mobility.
SmaI  (3189)
1 site
C C C G G G G G G C C C

SmaI can be used at 37°C for brief incubations.
TspMI  (3187)
1 site
C C C G G G G G G C C C
XmaI  (3187)
1 site
C C C G G G G G G C C C

Cleavage may be enhanced when more than one copy of the XmaI recognition sequence is present.
PstI  (3185)
1 site
C T G C A G G A C G T C
HindIII  (3163)
1 site
A A G C T T T T C G A A
SalI  (3148)
1 site
G T C G A C C A G C T G
AbsI  (3142)
1 site
C C T C G A G G G G A G C T C C
PaeR7I  (3142)
1 site
C T C G A G G A G C T C

PaeR7I does not recognize the sequence CTCTCGAG.
PspXI  (3142)
1 site
V C T C G A G B B G A G C T C V
XhoI  (3142)
1 site
C T C G A G G A G C T C
ApaI  (3137)
1 site
G G G C C C C C C G G G

ApaI can be used between 25°C and 37°C.
PspOMI  (3133)
1 site
G G G C C C C C C G G G
NaeI  (2801)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NaeI recognition sequence.
NgoMIV  (2799)
1 site
G C C G G C C G G C C G

Efficient cleavage requires at least two copies of the NgoMIV recognition sequence.
PfoI  (46)
1 site
T C C N G G A A G G N C C T

Sticky ends from different PfoI sites may not be compatible.
PflFI  (192)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by PflFI may be hard to ligate.
Sticky ends from different PflFI sites may not be compatible.
Tth111I  (192)
1 site
G A C N N N G T C C T G N N N C A G

The 1-base overhangs produced by Tth111I may be hard to ligate.
Sticky ends from different Tth111I sites may not be compatible.
BsrGI  (632)
1 site
T G T A C A A C A T G T

BsrGI is typically used at 37°C, but is even more active at 60°C.
BfuAI  (759)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BfuAI recognition sequence.
Sticky ends from different BfuAI sites may not be compatible.
BfuAI is typically used at 50°C, but is 50% active at 37°C.
BspMI  (759)
1 site
A C C T G C ( N ) 4 T G G A C G ( N ) 4 ( N ) 4

Efficient cleavage requires at least two copies of the BspMI recognition sequence.
Sticky ends from different BspMI sites may not be compatible.
XcmI  (1201)
1 site
C C A N N N N N N N N N T G G G G T N N N N N N N N N A C C

The 1-base overhangs produced by XcmI may be hard to ligate.
Sticky ends from different XcmI sites may not be compatible.
BbsI  (1301)
1 site
G A A G A C N N C T T C T G N N ( N ) 4

Sticky ends from different BbsI sites may not be compatible.
BbsI gradually loses activity when stored at -20°C.
AflII  (1438)
1 site
C T T A A G G A A T T C
AgeI  (1503)
1 site
A C C G G T T G G C C A
BstEII  (1718)
1 site
G G T N A C C C C A N T G G

Sticky ends from different BstEII sites may not be compatible.
BstEII is typically used at 60°C, but is 50% active at 37°C.
PpuMI  (1875)
1 site
R G G W C C Y Y C C W G G R

Sticky ends from different PpuMI sites may not be compatible.
HpaI  (2174)
1 site
G T T A A C C A A T T G
KasI  (2203)
1 site
G G C G C C C C G C G G
NarI  (2204)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the NarI recognition sequence.
SfoI  (2205)
1 site
G G C G C C C C G C G G
PluTI  (2207)
1 site
G G C G C C C C G C G G

Efficient cleavage requires at least two copies of the PluTI recognition sequence.
BsaAI  (2698)
1 site
Y A C G T R R T G C A Y
DraIII  (2698)
1 site
C A C N N N G T G G T G N N N C A C

Sticky ends from different DraIII sites may not be compatible.
BtgZI  (2699)
1 site
G C G A T G ( N ) 10 C G C T A C ( N ) 10 ( N ) 4

Sticky ends from different BtgZI sites may not be compatible.
After cleavage, BtgZI can remain bound to DNA and alter its electrophoretic mobility.
BtgZI is typically used at 60°C, but is 75% active at 37°C.
LEU2
663 .. 1757  =  1095 bp
364 amino acids  =  39.0 kDa
Product: 3-isopropylmalate dehydrogenase, required for leucine biosynthesis
yeast auxotrophic marker
LEU2
663 .. 1757  =  1095 bp
364 amino acids  =  39.0 kDa
Product: 3-isopropylmalate dehydrogenase, required for leucine biosynthesis
yeast auxotrophic marker
AmpR
4447 .. 5307  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 2:  
   4447 .. 5238  =  792 bp
   263 amino acids  =  28.9 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
4447 .. 5307  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
   Segment 1:  signal sequence  
   5239 .. 5307  =  69 bp
   23 amino acids  =  2.6 kDa
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
AmpR
4447 .. 5307  =  861 bp
286 amino acids  =  31.6 kDa
2 segments
Product: β-lactamase
confers resistance to ampicillin, carbenicillin, and related antibiotics
ori
3688 .. 4276  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin of replication
ori
3688 .. 4276  =  589 bp
high-copy-number colE1/pMB1/pBR322/pUC origin of replication
lacZα
2712 .. 3290  =  579 bp
192 amino acids  =  20.6 kDa
Product: LacZα fragment of β-galactosidase
lacZα
2712 .. 3290  =  579 bp
192 amino acids  =  20.6 kDa
Product: LacZα fragment of β-galactosidase
LEU2 promoter
1770 .. 2174  =  405 bp
LEU2 promoter
1770 .. 2174  =  405 bp
AmpR promoter
5308 .. 5412  =  105 bp
AmpR promoter
5308 .. 5412  =  105 bp
lac promoter
3334 .. 3364  =  31 bp
3 segments
   Segment 3:  -10  
   3334 .. 3340  =  7 bp
promoter for the E. coli lac operon
lac promoter
3334 .. 3364  =  31 bp
3 segments
   Segment 2:  
   3341 .. 3358  =  18 bp
promoter for the E. coli lac operon
lac promoter
3334 .. 3364  =  31 bp
3 segments
   Segment 1:  -35  
   3359 .. 3364  =  6 bp
promoter for the E. coli lac operon
lac promoter
3334 .. 3364  =  31 bp
3 segments
promoter for the E. coli lac operon
lac operator
3310 .. 3326  =  17 bp
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
lac operator
3310 .. 3326  =  17 bp
The lac repressor binds to the lac operator to inhibit transcription in E. coli. This inhibition can be relieved by adding lactose or isopropyl-β-D-thiogalactopyranoside (IPTG).
f1 ori
2474 .. 2929  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
f1 ori
2474 .. 2929  =  456 bp
f1 bacteriophage origin of replication; arrow indicates direction of (+) strand synthesis
MCS
3127 .. 3234  =  108 bp
pBluescript multiple cloning site
MCS
3127 .. 3234  =  108 bp
pBluescript multiple cloning site
T7 promoter
3100 .. 3118  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T7 promoter
3100 .. 3118  =  19 bp
promoter for bacteriophage T7 RNA polymerase
T3 promoter
3247 .. 3265  =  19 bp
promoter for bacteriophage T3 RNA polymerase
T3 promoter
3247 .. 3265  =  19 bp
promoter for bacteriophage T3 RNA polymerase
M13 fwd
3074 .. 3090  =  17 bp
common sequencing primer, one of multiple similar variants
M13 fwd
3074 .. 3090  =  17 bp
common sequencing primer, one of multiple similar variants
M13 rev
3286 .. 3302  =  17 bp
common sequencing primer, one of multiple similar variants
M13 rev
3286 .. 3302  =  17 bp
common sequencing primer, one of multiple similar variants
ORF:  784 .. 1095  =  312 bp
ORF:  103 amino acids  =  11.1 kDa
ORF:  4577 .. 4843  =  267 bp
ORF:  88 amino acids  =  9.2 kDa
ORF:  663 .. 1769  =  1107 bp
ORF:  368 amino acids  =  39.4 kDa
ORF:  2712 .. 3290  =  579 bp
ORF:  192 amino acids  =  20.6 kDa
ORF:  4447 .. 5307  =  861 bp
ORF:  286 amino acids  =  31.6 kDa
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